A tentative schedule of things we will do over the first few weeks. ------ Old 2012 Schedule
Journal Club Schedule: Journal Club Summ13 Researcher's Names:
Tony, Alyssa, Grace, Young, Jacky, Kevin, Katherine, Kavya, Marianna, Melissa, Nicolet, Priya, Ramiro, Vicky, Oscar, Jason, Betty, Brian, Gabe, Salil, Stephanie, Ariel
Renee, Serena, Will, Madeline, til ~July 15th, Ashlee -til July 3rd, Anita, Laraib, Karuna - til June 28th, Brendan,
NOTE - see very bottom of page for previous weeks' schedules and things to catch up on that you may have missed.
RESEARCH POSTER WORKSHOP SERIES: Tues, July 23rd: Research Poster Design Workshop FAC 328 3-4 PM
Tues, July 30th: Research Poster Design Workshop FAC 328 3-4 PM
Wed, July 31st: Research Poster Presentation Workshop FAC 328 3-4 PM
Tues, Aug 6th: Research Poster Presentation Workshop FAC 328 3-4 PM
Thurs Aug 8th: Poster Session WEL Grand Hall 1-3 PM
This Week
People to update Target page: People to update Wikispaces Research Pages:
MONDAY 29-Jul
Weekly Group Meeting @ 1:00 PM inGEA 403 (Gearing Hall)
3:00 PM - Research Poster Design Wokshop FAC 328
-- Good preparation for the poster you will generate in the Spring for the Undergraduate Research Forum OR if you want to attend a travel conference (e.g. ABRCMS or NCUR)
TUESDAY 30-Jul
WEDNESDAY 31-Jul
Last official day in the lab
Check your hours on the Hours Chart Be sure you have enough towards you 160.
EVERYONE:
Staff Stuff:
Label new reagents and supplies
Dilute new 100 bp ladder in Blue Juice + TE
test all tubes of [[#|Taq Polymerase]] in PCR
Target Discovery choices --> order primer sets
ToDo List:
Lab Notebook Checks
Protocol cost analysis (Transformation, cloning)
Walk through protein expression (use current mentor's enzymes)
Joey & Christina's Francisella HAP enzyme
Sadhana's M. tuberculosis enzyme
Joshua's M. tuberculosis enzyme
J.C.'s YopH
make and run SDS-Page protein gels
load and run SDS-Page protein gels
Lab Group Meeting/Journal Club (in
Final Summer Research Presentations (upload to Google Docs by Sunday night)
2-3 slides, ~ 7 minutes
What your disease and protein target are (since the rest of the group doesn't know exactly what you are working on)
Specifics about size of gene, GC% content, etc.
Size of protein, is there a structure in the PDB?
What you set out to do
Where you have gotten so far (show gel pics, images, graphs or relevant data)
Challenges, failures, side-steps
Future Directions (i.e. Fall semester and beyond)
Social ???
(Remember to work on your 1 page (~500) word summary of your research. No figures needed.
- Motivation, What you did, What you have left to do on your project. Reflect on what you have gotten out of your summer research and if it was what you expected.
- submit to Dr. B before Sept. 1st. Then I will proof and send to Dr. Simmons.
.
Last Day of VDS Summer Research
expression, purification, characterization
test specific activity of your enzyme for lot to lot consistency
Research Presentation Schedule:
Date: Name
Instructions: prepare 3-5 Powerpoint slides to present your work to date
Intro (talk about your disease and what your protein does), Objectives, Methods& Results, Future Directions
OLD STUFF (2012 Summer) Dr. B NOTES
PCRs to do succesfully before you can move on to Target cloning:
ProtocolPCRforREcloningGREEN.xls OR ProtocolPCRforREcloningRED.xls
ProtocolPCR_pLICsequencing.xls
Veteran VDSers:
Pick up target tail primers (Leader and Laggers) we ordered 2 days ago (to get in MBB building)
ProtocolPrimerDilutionVDS_Summ12.docx
Make your Oligo Mixfor your target (first step in Overlap PCR protocol) using PLATE of overlap primers - VERY important to not cross-contaminate your primers!!
ProtocolPCR_PrimerOverlapVDS_Summ12.docx
takes A LONG time to thaw primers in box.
Transformation Efficiency: pmCherry pGFP
Veteran VDSers:
Make Kan/Sucrose LB Agar plates for your target cloning
ProtocolPCRforREcloningGREEN.xls OR ProtocolPCRforREcloningRED.xls
After your Primary and Secondary Overlap PCR work, go to : ProtocolpNIC-Bsa4_cloningVDS_Summ12.doc
make SDS-Page protein gels for characterization step
Midiprep on pNIC-Bsa4 :
Midiprep on pUC19 : ??
Transformation of plasmids to make stocks - pUC19
12:30, 1:00 - all day: protein expression (Max , Kaarthik , Urvashi , Janice , Rishi, Aldo (observe) )
-- we will start a little later than we normally would but we will use an abbreviated induction phase (~ 3 hrs) so that we can finish in a more reasonable time.
(NOT YET) ProtocolBacterialGrowthCurve_VDS_Summ12.xlsx - test (needs to be modified a little)
Each Person:
one flask at 30degC in water bath shaker
one flask at 37deg in air incubator
Measure OD every few hours over ~ 8hrs time span until OD reaches 1.000 at 600nm
Make LB and Autoclave with flasks that you will use.
------- put 500ml into in 2x 2L flasks
------ put 50 ml into 4 x 250 ml flasks
Protein expression
Monitor log phase growth in Excel spreadsheet graph
make SDS-Page protein gels for characterization step
EVERYONE: determine gene inside pNIC-Bsa4 by checking your pLIC sequencing that you sent to the core the other week
EVERYONE THAT HAS DONE MIDI-Prep (successfully): Have you submitted your sample to sequencing? Does it turn out to be the right plasmid?
~ 10:00 Prep Starter Culture (grow until ~ 8hrs to OD600 = 1 )
Post your class schedule to GDOcs - Summer13
Create your own folder in Google Docs under the folder: /StudentFolders
MONDAY 3-Jun
2:00 - PAI 2.14 First meeting
1st Journal Club (please read the journal club article prior to meeting so that we can discuss it in groups)
Zolli-Juran, M.; Cechetto, J. D.; Hartlen, R.; Daigle, D. M.; Brown, E. D., High throughput screening identifies novel inhibitors of Escherichia coli dihydrofolate reductase that are competitive with dihydrofolate. Bioorganic & Medicinal Chemistry Letters 2003, 13 (15), 2493-2496. Zolli_JuranDHFRbacteriaHTSBioorgMedChemLett2003.pdf
Lab Safety training
- New guys - walk through with Dr. B
Add content to YOUR research page Research Pages 2013
Put a line and 'Week 1' Below that add your 'result' image for the week with brief commentary
This page will run in reverse chronological order - i.e. Newest stuff on top
Order Overlap Primers for class
Journal article for next week is posted on Journal Club page.
SATURDAY 8-Jun
SUNDAY 9-Jun
Week 2
MONDAY 10-Jun
Safety training check.
Weekly Group Meeting @ 1:00 PM inGEA 100 (Gearing Hall)
Target Discovery selections
Protocol Primer Design OverlapAssembly
Order Overlap Primers for class
1pm start agarose gels of last week's 1st PCR (for the people not doing MidiPrep - e.g. Katherine, Nicolet, Marianna, Kevin, Oscar, Jason, Serena, Kavya, Jacky)
PCR ? - Laraib, Betty
Submit to DNA sequencing - Laraib, Betty, Oscar, Jason
Nanodrop a plasmid - Laraib, Betty, Oscar, Jason
Do overnight cultures of plasmids - Oscar, Jason, Laraib, Betty
Make 500 ml of LB
New guys do Lab 1: Buffers & Solutions
On Your Targets Page:
Add Link to DNA Works output text file
Add substrate cost, quantity, and catalog numbers (and supplier) for enzyme assay reagents
Add image of BRENDA reaction mechanism (screenshot it)
Fill out Travel Authorization Forms for field trips.
-- see GoogleDocs/Misc/TravelForms
-- print out and complete - put in Dr. B's box in lab (pigeon hole)
[ACTIONREQUIRED]: Will/Ramiro please put your DNAWorks output file into the "DNAWorks_Output" folder in vdsclass@gmail.com/Misc/Primers.
Grace/Young, Priya/Vicky, Jacky/Kavya, and Serena please upload your DNA oligonucleotide sequences onto the spreadsheet titled "PrimerPlateOrderTemplate_GDocs_Summer2013".
THURSDAY 13-Jun
Repeat 1st PCR if failed (why did it fail?)
Then:
ProtocolPCRforREcloningGREEN.xls OR ProtocolPCRforREcloningRED.xls
Analyze DNA Sequence Exercise as a group?
FRIDAY 14-Jun
11 am - 2pm Optional time to catch up on stuff. - Daniel mentoring
Print Journal Club article for you and your friends
Make LB for next week (500ml per person)
Agarose gel checks
ProtocolPCRforREcloningGREEN.xls OR ProtocolPCRforREcloningRED.xls
Below that add your 'result' image for the week with brief commentary
This page will run in reverse chronological order - i.e. Newest stuff on top
Lab Notebook check - should have everything up to date.
SATURDAY 15-Jun
-- sleep all day SUNDAY 16-Jun
-- watch TV all day
WEEK 3
When not doing protein expression - can work on PCR's Red/Green, and then PCR pLIC sequencing,
ProtocolPCRforREcloningRED.xls OR ProtocolPCRforREcloningGREEN.xls (if fail, redo)
ProtocolPCR_pLICsequencing.xls (if fail, redo)
Grow up your own batch of pNIC-Bsa4 (160ml) overnight and spin down to pellets the next day, save, and midi-prep later. (if fail, redo - because you need it for cloning)
MONDAY 17-Jun
Those who need to update their Wikispaces page: Young, Karuna, Renee, Kevin, Madeline, Manuel, Nicolet, William
Verify your Safety training is all present.
1:00 - 4 people (Grace, Katherine, Young, Nicolet) start transformation of YopH and FtHAP into BL21(DE3) cells (instead of DH5alpha)
---- plasmids should be in 'Plasmids' box or the 'Final Plasmids' box
---- share your tube of competent bacterial cells BL21(DE3) with another groups - i.e. only use 1/2 of the tube for yours
Grace & Kavya - YopH in pNIC-Bsa4
Katherine & Jacky - FtHAP in pNIC-Bsa4 Young - YopH in pNIC-Bsa4
Nicolet & Kevin - FtHAP in pNIC-Bsa4
OTHERS: Start one of the PCRs listed above
OR Make LB for expression if you haven't already done so (500ml per person)
AND/OR Prepare your flasks for overnight culture for protein expression (2x 50ml OR 2 x 100 ml of LB with Antibiotic) - Alyssa, Melissa, Vicky, Priya, Tony, Renee, Ramiro, Will
Label your flasks
AND/OR Check DNA sequence of your MidiPrepped samples
-- once the plasmid is verified - move it to the regular 'plasmids' box (and but the other half in the top loading freezer in the 'plasmids' box)
-- Update plasmid info on the Google 'Plasmid Concentrations' spreadsheet:
---------https://docs.google.com/spreadsheet/ccc?key=0AoO2KqKh2q_-dENvNGJEdW9qdUxQZmFYeTVoeXA0SUE&usp=sharing
Weekly Group Meeting@ 1:00 PMCHANGED TO @ 3:00 PM inGEA 100 (Gearing Hall) - unless we can't get this room. & Journal Club - see Journal Club page for paper
Daniel - create a 'Plasmids' box in the top loading freezer (already have one for the other freezer)
Analyze DNA sequence exercise (as a group) - in WEL computer lab
5: 00 - Overnight growth of pGBR22 2x 80 ml of LB, Midiprep tomorrow (Select 2 people) ? Betty & Laraib?
5:30 - start overnight cultures (only those people who are expressing tomorrow) (Alyssa, Melissa, Vicky, Priya, Tony, Renee, Ramiro, Will)
TUESDAY 18-Jun 10 am - Larger Scale Protein Expressions (work in pairs)
ProtocolProteinExpression_VDS_Summ13_v1.doc
USE THIS PROTOCOL TOO: ProtocolBacterialGrowthCurve_VDS_Summ13.xlsx
Alyssa (Not Mon or Wed) & Melissa - FabV in pNIC-Bsa4PfDXR in pNic-BSSA4 Vicky & Priya - PfDXR in pNic-BSSA4 Tony & Renee (has class til 11:30) - FabI in pNIC-Bsa4PfDXR in pNic-BSSA4 Ramiro (Tues best, Monday ok, Not Wed, Thurs?) & Will - FrTUHP in pnic-BSA4 PfDXR in pNic-BSSA4
1:00 - everyone else come in to catch up on PCR's, agarose gel checks
Mentors (i.e. Michael) : During the break in expression
- make Resuspension buffer (~10ml per person) - see GDocs/Misc/Solutions&Dilutions or see protocol for protein expression
- make more 1 M IPTG
Transfer your transformation plate to 4degC fridge (label it correctly, and parafilm) - only these VDSers:
Grace & Kavya - YopH in pNIC-Bsa4
Katherine & Jacky - FtHAP in pNIC-Bsa4 Young - YopH in pNIC-Bsa4
Nicolet & Kevin - FtHAP in pNIC-Bsa4
5:30 - start overnight cultures (only those people who are expressing tomorrow) - Grace, Kavya, Katherine, Jacky, Madeline, Brendan, Kevin, Marianna, Serena, Jason, Oscar
WEDNESDAY 19-Jun 10 am - Larger Scale Protein Expressions (work in pairs)
ProtocolProteinExpression_VDS_Summ13_v1.doc
USE THIS PROTOCOL TOO: ProtocolBacterialGrowthCurve_VDS_Summ13.xlsx Grace (Not Tuesday) & Kavya - (Kavya has every day til 11:30 am (could skip on any day but Monday) - prefers Wed) - YopH in pNIC-Bsa4 Katherine (but Tues afternoon class) & Jacky (not Mon - Tuesday is ok, Wednesday better) - FtHAP in pNIC-Bsa4 Madeline & Brendan - any day if advance notice..... - YopH in pNIC-Bsa4 Kevin(Kevin has class till 11:15) & Marianna - has class 8:30 - 11:30 (Tues is class all day) - FtHAP in pNIC-Bsa4 Serena FF @ 11 on Th - YopH in pNIC-Bsa4 Jason (10am free) & Oscar FtHap in pNIC-Bsa4
1:00 - people from yesterday - resuspend cell pellets in 10ml of buffer and store in conical tubes in -80degC
5:30 - start overnight cultures (only those people who are expressing tomorrow)
THURSDAY 20-Jun (Dr. B out of town) - Daniel and Co. are in charge Firefighter Training ? - who is doing this....
Thurs @ 9:00 - 10: Karuna, Ashlee, Alyssa, Renee, Priya
Thurs @ 11- 12: Melissa, Tony, Oscar, Serena, Kavya, Anita, Marianna, Katherine
10 am - Larger Scale Protein Expressions (work in pairs)
ProtocolProteinExpression_VDS_Summ13_v1.doc
USE THIS PROTOCOL TOO: ProtocolBacterialGrowthCurve_VDS_Summ13.xlsx
Young - FtHAP in pNIC-Bsa4
Ashlee & Karuna (karuna has class til 11:30) - FabI in pNIC-Bsa4
Nicolet & Anita - YopH in pNIC-Bsa4
Betty & Laraib - (Laraib class 10-11:50 each day) - PfDXR in pNic-BSSA4
1:00 - people from yesterday - resuspend cell pellets in 10ml of buffer and store in conical tubes in -80degC
EVERYONE: complete and place your Release form and Medical Waiver in Dr. B's 'pigeon hole' (the one that says RESERVED over by the lab notebooks)
PROTEIN GROUPS (from last week) Group A Alyssa & MelissaPfDXR in pNic-BSSA4 Vicky & Priya - PfDXR in pNic-BSSA4 Tony & ReneePfDXR in pNic-BSSA4 Ramiro & WillPfDXR in pNic-BSSA4
Group B Grace & Kavya - YopH in pNIC-Bsa4
Madeline & Brendan YopH in pNIC-Bsa4 Kevin & Jacky- FtHAP in pNIC-Bsa4 Serena YopH in pNIC-Bsa4 Jason & Oscar FtHap in pNIC-Bsa4
Group C Young - FtHAP in pNIC-Bsa4 Ashlee & Karuna FabI in pNIC-Bsa4 Nicolet & Anita YopH in pNIC-Bsa4 Betty & Laraib PfDXR in pNic-BSSA4
Katherine & Marianna FtHAP in pNIC-Bsa4
MONDAY 24-Jun Weekly Group Meeting @ 1:00 PM inGEA 100 (Gearing Hall) & Journal Club - see Journal Club page for paper
Analyze DNA sequence exercise (as a group) - in WEL computer lab
EVERYONE: complete and place your Release form and Medical Waiver in Dr. B's 'pigeon hole' (the one that says RESERVED over by the lab notebooks)
TUESDAY 25-Jun
1:00 Group A:Alyssa & MelissaPfDXR in pNic-BSSA4, Vicky & Priya - PfDXR in pNic-BSSA4, Tony & ReneePfDXR in pNic-BSSA4, Ramiro & WillPfDXR in pNic-BSSA4
Make Homemade Protein (SDS-PAGE) Gels
Sonication & spin down
Group B:Grace & Kavya - YopH in pNIC-Bsa4, , Madeline & Brendan YopH in pNIC-Bsa4, Kevin & Jacky- FtHAP in pNIC-Bsa4, Serena YopH in pNIC-Bsa4, Jason & Oscar FtHap in pNIC-Bsa4
Sonication & spin down (maybe on to purification?)
Group C: Young - FtHAP in pNIC-Bsa4, Ashlee & Karuna FabI in pNIC-Bsa4, Nicolet & Anita YopH in pNIC-Bsa4, Betty & Laraib PfDXR in pNic-BSSA4
Katherine & Marianna FtHAP in pNIC-Bsa4
RE digest (during incubation period - make gels)
5:00 - After RE digest, people from last Thursday's expression (Group C) - resuspend cell pellets in 10ml of buffer and store in conical tubes in -80degC (20-30 minutes)
WEDNESDAY 26-Jun
11:00 AM
Group A:Alyssa & MelissaPfDXR in pNic-BSSA4, Vicky & Priya - PfDXR in pNic-BSSA4, Tony & ReneePfDXR in pNic-BSSA4,
Sonication & Spin Down & Purification & run SDS-Page gels
1:00 PM Group B:Grace & Kavya - YopH in pNIC-Bsa4, Madeline & Brendan YopH in pNIC-Bsa4, Kevin & Jacky- FtHAP in pNIC-Bsa4, Serena YopH in pNIC-Bsa4, Jason & Oscar FtHap in pNIC-Bsa4 RE digest (during incubation period - make gels)------ Change to Purification RE digest (during incubation period -
Make Homemade Protein (SDS-PAGE) Gels
Group C: Young - FtHAP in pNIC-Bsa4, Ashlee & Karuna FabI in pNIC-Bsa4, Nicolet & Anita YopH in pNIC-Bsa4, Betty & Laraib PfDXR in pNic-BSSA4, Katherine & Marianna FtHAP in pNIC-Bsa4,, and Ramiro & Will PfDXR in pNic-BSSA4
Sonication& Spin Down), purification
THURSDAY 27-Jun
10:00 AM Group A:Alyssa & MelissaPfDXR in pNic-BSSA4, Vicky & Priya - PfDXR in pNic-BSSA4, Tony & ReneePfDXR in pNic-BSSA4, Ramiro & WillPfDXR in pNic-BSSA4
run gels, stain, destain
Group C: Young - FtHAP in pNIC-Bsa4, Ashlee & Karuna FabI in pNIC-Bsa4, Nicolet & Anita YopH in pNIC-Bsa4, Betty & Laraib PfDXR in pNic-BSSA4, Katherine & Marianna FtHAP in pNIC-Bsa4, purification
For everyone else - Open time - come in to work on stuff (PCR's, gels, Wikispaces updating, Notebooks, etc.)
12:30 PM - NOTE TIME CHANGE - we need to be early enough to catch the bus. EVERYONE: Gather together to get on bus to go to Pickle Research Campus for the TACC Tour (starts at 2pm there)
Ramiro & Will PfDXR in pNic-BSSA4 - add Ni-NTA resing (only 0.5 ml) overnight
WEEK 4
FRIDAY 28-Jun -
11:00 AM
finish purifications, nanodrop, enter data to Google Spreadsheet (Protein Spectrophotometer Calcs)
Run gels (Protein gels), stain for 1 hour, destain overnight
Dry gels if already de-stained
Make protein gels (if we have enough combs)
RE digest if not doing protein stuff
Grow up your own pNIC-Bsa4, mdidiprep, sequence verify it, store in your own box
Cut your pNIC-Bsa4 to prepare the vector for cloning, save in -20degC in your box
WEDNESDAY 3-Jul
11:00 AM - mentors on duty
Dilute tail primers
submit pNIC-Bsa4 to sequencing, to verify plasmid & determine what gene is in the middle
THURSDAY 4-Jul
OFF - celebrate and shoot off some fireworks
FRIDAY 5-Jul
11:00 AM - mentors on duty
We'll have open lab time (with mentors)- for people to catch up on stuff but not an organized activity
??
1:00 Re-run VS4 docking with Histidines protonated (Group 2)
??
1:00 FPLC of PfDXR - for those that expressed it.
submit pNIC-Bsa4 to sequencing, to verify plasmid & determine what gene is in the middle
SATURDAY 6-Jul
Do people need more hours? - If so we can have lab times on Saturday. Let Dr. B know via email or in person (or carrier pigeon).
SUNDAY 7-Jul
Invent a new language
6th Week
PROTEN GROUPS FROM OTHER WEEK: Group A:Alyssa & MelissaPfDXR in pNic-BSSA4, Vicky & Priya - PfDXR in pNic-BSSA4, Tony & ReneePfDXR in pNic-BSSA4, Ramiro & WillPfDXR in pNic-BSSA4 Group B:Grace & Kavya - YopH in pNIC-Bsa4, , Madeline & Brendan YopH in pNIC-Bsa4, Kevin & Jacky- FtHAP in pNIC-Bsa4, Serena YopH in pNIC-Bsa4, Jason & Oscar FtHap in pNIC-Bsa4 Group C: Young - FtHAP in pNIC-Bsa4, Ashlee FabI in pNIC-Bsa4, Nicolet & Anita YopH in pNIC-Bsa4, Betty & Laraib PfDXR in pNic-BSSA4
Katherine & Marianna FtHAP in pNIC-Bsa4 MONDAY 8-Jul
Weekly Group Meeting @ 1:00 PM inGEA 403 (Gearing Hall) -NOTE THE ROOM CHANGE!!!!!! Same building
& Journal Club - see Journal Club page for paper
After class - PfDXR group spins down sample (combine together) Alyssa & Melissa, Vicky & Priya,Tony & ReneePfDXR, Ramiro , Betty
4:00 - 5:15 - Naturally Obsessed documentary viewing in PAI 3.02 (with popcorn?)
TUESDAY 9-Jul
1:00 - FtHAP group spins down sample (combine together)
FtHAP group: Kevin & Jacky, Jason & Oscar, Katherine,
1 PM - FPLC for PfDXR group Alyssa & Melissa, Vicky & Priya - PfDXR, Tony & ReneePfDXR, RamiroPfDXR
Learn Virtual Screening:
Oscar, Jason, Stephanie, Brian, Gabe, Betty, Salil
4:00 - FPLC people collect fractions of protein and spin down again, Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
? Store in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC)
WEDNESDAY 10-Jul
1:00 - YopH group spins down sample (combine together)
YopH group: Grace & Kavya, Brendan, Serena, Nicolet, Young
1 PM - FPLC for FtHAP group
FtHAP group: Kevin & Jacky, Jason & Oscar, Katherine
2 - 4pm: Middle Schoolers (Shadow a Scientist) - Alyssa & Tony
4:00 - FPLC people collect fractions of protein and sping down again, Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
? Store in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC)
THURSDAY 11-Jul
1:30 or 2:00 PM - FPLC for YopH group
YopH group: Grace & Kavya, Serena, Young
1:00
PfDXR group - Alyssa & Melissa, Vicky & Priya, Tony & Renee, Ramiro - with ANDREW
YopH (i.e. PTP1b) group -Kevin, Jason & Oscar, Katherine - with DANIEL
Spin down FPLC'ed sample from the other day (to about 2 ml)
Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
Go get liquid nitrogen (with a mentor!)
Store in protein in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC) 1:00 - FtHAP group spins down sample (combine together)
3:00 Re-run VS4 docking with Histidines protonated (Group 3):
Grace, Young, Kavya, Ariel
Virtual Screening 1 (Part 2) - using GOLD: Oscar, Jason, Betty, Stephanie, Salil, Gabe, Brian
~ 4:00 - FPLC people collect fractions of protein and sping down again, Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
?
Dr. B in a workshop all day Daniel & Michael are in charge (scary!)
1:15 PM - TACC VisLab Tour (on campus in ACES building) - meet in our lab first and walk over
After TOUR:
YopH group: Grace & Kavya, Serena, Young
Spin down FPLC'ed sample from the other day (to about 2 ml)
Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
Go get liquid nitrogen (with a mentor!)
Store in protein in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC) 1:00 - FtHAP group spins down sample (combine together)
Grow up your own pNIC-Bsa4, midiprep, sequence verify it, store in your own box
after submitting pNIC-Bsa4 to sequencing - determine what gene is in the middle
Cut your pNIC-Bsa4 to prepare the vector for cloning (one of the 1st steps in the cloning protocol), save in -20degC in your box
Continue Overlap PCR's
Begin cloning after successful PCR's
Target group meetings with Dr. B
Everyone make two (2) Kan+Sucrose LB Agar plates - we have lots (thank to Jacky, Oscar and Priya!) , but I want you to know how to make them.
-- make 20% (w/v) sucrose solution, sterile filter it ........ w/v = weight to volume
-- make LB agar plates as usual, autoclave
-- after cooled, add Kan and Sucrose (final conc of Sucrose should be 5% (w/v)) - NOTE: don't want to autoclave Kan or Sucrose
GOLD on your target - validation library
ICM run
VINA run, windows box only
MONDAY 15-Jul
Weekly Group Meeting @ 1:00 PM inGEA 403 (Gearing Hall) -NOTE THE ROOM CHANGE!!!!!! Same building
& Journal Club - see Journal Club page for paper
2:30 - Session to update Wikispaces Target pages
Nanodrop protein, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
Go get liquid nitrogen (with a mentor!)
Store in protein in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC) TUESDAY 16-Jul
WEDNESDAY 17-Jul
THURSDAY 18-Jul
People to update Target page:Nicolet, Grace & Young, Vicky & Kavya, Jacky & Kevin
People to update Wikispaces Research Pages: Grace, Jacky, Jason, Katherine, Nicolet, Ramiro,
Send your research presentation slides to Dr. B:Tony, Katherine V., Alyssa & Stephanie, Grace & Young
FRIDAY 19-Jul
Check your hours on the Hours Chart Be sure you have enough towards you 160.
EVERYONE:
Below that add your 'result' image for the week with brief commentary
This page will run in reverse chronological order - i.e. Newest stuff on top
Lab Notebook check - should have everything up to date.
SATURDAY 20-Jul
SUNDAY 21-Jul
MONDAY 22-Jul
Weekly Group Meeting @ 1:00 PM inGEA 403 (Gearing Hall)
& Journal Club - see Journal Club page for paper
TUESDAY 23-Jul
Jacky, Kevin, Kavya, Vicky
grow up some pUC19
buy some Sma1 or HincII from ICMB Core Facility Store. Get NEB version.
Midi prep pUC19
make Xgal/IPTG plates with Ampicillin resistance
make some LB too (500ml)
Tues, July 23rd: Research Poster Design Workshop FAC 328 3-4 PM
- optional. Good preparation for the poster you will generate in the Spring for the Undergraduate Research Forum
WEDNESDAY 24-Jul
Send your research presentation slides to Dr. B: Nicolet, Ramiro, Jacky & Kevin, Melissa & Jason
THURSDAY 25-Jul
HSers (Brian, Gabe, Betty, Salil)
- work on Purchasing budget for reagents. In GoogleDrive/Misc/Purchasing
- examples of past orders are on second sheet in the file.
- copy and past a past order to the first sheet and then modify it to reflect what you used - Thx. - Dr. B
Then do VS3
Grace & Young - meet with Dr. B about journal paper
Send slides to Dr. B - Nicolet, Ramiro, Jacky & Kevin,
Journal Club Schedule:
Journal Club Summ13
Researcher's Names:
Tony, Alyssa, Grace, Young, Jacky, Kevin, Katherine, Kavya, Marianna, Melissa, Nicolet, Priya, Ramiro, Vicky, Oscar, Jason, Betty, Brian, Gabe, Salil, Stephanie, Ariel
Renee, Serena, Will, Madeline, til ~July 15th, Ashlee -til July 3rd, Anita, Laraib, Karuna - til June 28th, Brendan,
NOTE - see very bottom of page for previous weeks' schedules and things to catch up on that you may have missed.
SEE THE ANNOUNCEMENTS PAGE FOR SUMMER SOCIAL FORM
Research Presentations Schedule
Monday 15th: Priya & Oscar & Betty, Kavya & VickyMonday 22nd: Tony, Katherine V., Alyssa & Stephanie, Grace & Young
Monday 29th: Nicolet, Ramiro, Jacky & Kevin, Melissa & Jason
RESEARCH POSTER WORKSHOP SERIES:
Tues, July 23rd: Research Poster Design Workshop FAC 328 3-4 PM
Tues, July 30th: Research Poster Design Workshop FAC 328 3-4 PM
Wed, July 31st: Research Poster Presentation Workshop FAC 328 3-4 PM
Tues, Aug 6th: Research Poster Presentation Workshop FAC 328 3-4 PM
Thurs Aug 8th: Poster Session WEL Grand Hall 1-3 PM
This Week
People to update Target page:People to update Wikispaces Research Pages:
MONDAY 29-Jul
Weekly Group Meeting @ 1:00 PM in GEA 403 (Gearing Hall)
& Journal Club - see Journal Club page for paperhttp://vdsstream.wikispaces.com/Journal+Club+Summer+13
3:00 PM - Research Poster Design Wokshop FAC 328
-- Good preparation for the poster you will generate in the Spring for the Undergraduate Research Forum OR if you want to attend a travel conference (e.g. ABRCMS or NCUR)
TUESDAY 30-Jul
WEDNESDAY 31-Jul
Last official day in the lab
Check your hours on the Hours Chart Be sure you have enough towards you 160.
EVERYONE:
Finish up your Lab Notebook and leave it in your pigeon hole
THURSDAY & FRIDAY - optional days. We will have mentors in the lab.
August 20th - 2 page summer reports due. Essentially a progress report/research update for the summer.
Next Week
nothing - empty your brain.Naturally Obsessed Form - when to watch this documentary- Please fill out this form....
https://docs.google.com/spreadsheet/viewform?formkey=dGdJaEQ5cUtEQm1uaXJ5LVNNdUR4NkE6MQ
TACC Tour at VisLab in ACES building ON CAMPUS - @
(meet in lab before at to walk over)Fill out - Medical Waiver
Fill out - Liability form
TACC Tour - @
(meet in lab before - then we'll catch bus up to Pickle campus)GASF (Genome Sequencing and Analysis Facility) Tour in MBB 4.102
Paper to read before the tour: NGS_Mardis.pdfAlgae Lab Tour: Meet in PAI 2.14 - then will walk over to BIO 320
Middle Schoolers visit lab
Cool Stuff We May Not Get To:
Library generation and ligand prep for virtual
Homology modelling
Screening with ICM
DSF (Differential Scanning Fluorimetry) assays
qPCR
Crystallization trials
Staff Stuff:
Label new reagents and supplies
Dilute new 100 bp ladder in Blue Juice + TE
test all tubes of [[#|Taq Polymerase]] in PCR
Target Discovery choices --> order primer sets
ToDo List:
Lab Notebook Checks
Protocol cost analysis (Transformation, cloning)
Walk through protein expression (use current mentor's enzymes)
make and run SDS-Page protein gels
load and run SDS-Page protein gels
Lab Group Meeting/Journal Club (in
Final Summer Research Presentations (upload to Google Docs by Sunday night)
Social ???
(Remember to work on your 1 page (~500) word summary of your research. No figures needed.
- Motivation, What you did, What you have left to do on your project. Reflect on what you have gotten out of your summer research and if it was what you expected.
- submit to Dr. B before Sept. 1st. Then I will proof and send to Dr. Simmons.
.
Last Day of VDS Summer Research
expression, purification, characterization
Research Presentation Schedule:
Date: Name
Instructions: prepare 3-5 Powerpoint slides to present your work to date
OLD STUFF (2012 Summer)
Dr. B NOTES
PCRs to do succesfully before you can move on to Target cloning:
ProtocolPCRforREcloningGREEN.xls OR ProtocolPCRforREcloningRED.xls
ProtocolPCR_pLICsequencing.xls
Veteran VDSers:
Transformation Efficiency:
pmCherry
pGFP
Veteran VDSers:
ProtocolMidiPrepKit_HiSpeed_VDS.doc on
(NOT YET) ProtocolBacterialGrowthCurve_VDS_Summ12.xlsx - test (needs to be modified a little)
Veteran VDSers:
After your Primary and Secondary Overlap PCR work, go to : ProtocolpNIC-Bsa4_cloningVDS_Summ12.doc
make SDS-Page protein gels for characterization step
Midiprep on pNIC-Bsa4 :
Midiprep on pUC19 : ??
Transformation of plasmids to make stocks - pUC19
12:30, 1:00 - all day: protein expression (Max , Kaarthik , Urvashi , Janice , Rishi, Aldo (observe) )
-- we will start a little later than we normally would but we will use an abbreviated induction phase (~ 3 hrs) so that we can finish in a more reasonable time.
(NOT YET) ProtocolBacterialGrowthCurve_VDS_Summ12.xlsx - test (needs to be modified a little)
Each Person:
- one flask at 30degC in water bath shaker
- one flask at 37deg in air incubator
Measure OD every few hours over ~ 8hrs time span until OD reaches 1.000 at 600nmMake LB and Autoclave with flasks that you will use.
------- put 500ml into in 2x 2L flasks
------ put 50 ml into 4 x 250 ml flasks
--ProtocolPrimerDilutionVDS_Summ12.docx
--pmCherry PCR
--pLIC PCR - ProtocolPCR_pLICsequencing.xls
--Overlap PCR of target
Make Protein Gels (Homemade SDS-Page) with
Protein Sonication
Protein expression
Monitor log phase growth in Excel spreadsheet graph
make SDS-Page protein gels for characterization step
EVERYONE: determine gene inside pNIC-Bsa4 by checking your pLIC sequencing that you sent to the core the other week
EVERYONE THAT HAS DONE MIDI-Prep (successfully): Have you submitted your sample to sequencing? Does it turn out to be the right plasmid?
~ 10:00 Prep Starter Culture (grow until ~ 8hrs to OD600 = 1 )
SATURDAY 29-May
SUNDAY 30-May
MONDAY 31-May
TUESDAY 1-Jun
WEDNESDAY 2-Jun
THURSDAY 3-Jun
FRIDAY 4-Jun
SATURDAY 5-Jun
SUNDAY 6-Jun
MONDAY 7-Jun
TUESDAY 8-Jun
WEDNESDAY 9-Jun
THURSDAY 10-Jun
FRIDAY 11-Jun
SATURDAY 12-Jun
SUNDAY 13-Jun
MONDAY 14-Jun
TUESDAY 15-Jun
WEDNESDAY 16-Jun
THURSDAY 17-Jun
FRIDAY 18-Jun
SATURDAY 19-Jun
SUNDAY 20-Jun
MONDAY 21-Jun
TUESDAY 22-Jun
WEDNESDAY 23-Jun
THURSDAY 24-Jun
FRIDAY 25-Jun
SATURDAY 26-Jun
SUNDAY 27-Jun
MONDAY 28-Jun
TUESDAY 29-Jun
WEDNESDAY 30-Jun
THURSDAY 1-Jul
FRIDAY 2-Jul
SATURDAY 3-Jul
SUNDAY 4-Jul
MONDAY 5-Jul
TUESDAY 6-Jul
WEDNESDAY 7-Jul
THURSDAY 8-Jul
FRIDAY 9-Jul
SATURDAY 10-Jul
SUNDAY 11-Jul
MONDAY 12-Jul
TUESDAY 13-Jul
WEDNESDAY 14-Jul
THURSDAY 15-Jul
FRIDAY 16-Jul
SATURDAY 17-Jul
SUNDAY 18-Jul
MONDAY 19-Jul
TUESDAY 20-Jul
WEDNESDAY 21-Jul
THURSDAY 22-Jul
FRIDAY 23-Jul
SATURDAY 24-Jul
SUNDAY 25-Jul
MONDAY 26-Jul
TUESDAY 27-Jul
WEDNESDAY 28-Jul
THURSDAY 29-Jul
FRIDAY 30-Jul
SATURDAY 31-Jul
SUNDAY 1-Aug
MONDAY 2-Aug
TUESDAY 3-Aug
WEDNESDAY 4-Aug
THURSDAY 5-Aug
FRIDAY 6-Aug
SATURDAY 7-Aug
SUNDAY 8-Aug
PREVIOUS WEEKS' SCHEDULES (for Summer 2013)
|| MONDAY || 3-Jun ||Week 1
Post your class schedule to GDOcs - Summer13Create your own folder in Google Docs under the folder: /StudentFolders
MONDAY 3-Jun
2:00 - PAI 2.14 First meeting
1st Journal Club (please read the journal club article prior to meeting so that we can discuss it in groups)
Zolli-Juran, M.; Cechetto, J. D.; Hartlen, R.; Daigle, D. M.; Brown, E. D., High throughput screening identifies novel inhibitors of Escherichia coli dihydrofolate reductase that are competitive with dihydrofolate. Bioorganic & Medicinal Chemistry Letters 2003, 13 (15), 2493-2496.
Zolli_JuranDHFRbacteriaHTSBioorgMedChemLett2003.pdf
Lab Safety training
- New guys - walk through with Dr. B
http://vdsstream.wikispaces.com/Required+Lab+Safety
Dilute some primers
'Learn how to submit to DNA sequencing' (starring Daniel et al.) - wet lab
TUESDAY 4-Jun
1:00 PM start time.......
Finish up yesterday's protocols...
New guys
- pipetting exercise and Buffers & Solutions (Lab 1 of Spring VDS)
Make LB and LB plates
ProtocolLBAgarPlatesVDS.doc
Choose a Plasmid to work with on these next 2 exercises(pGBR22 or pGFP or pmCherry or pNIC-Bsa4 or pUC19)
Several people: Transform a plasmid for Midi-prep (make lots of pGBR22 and pNIC-Bsa4, make a few pGFP and pmCherry and pUC19)
WEDNESDAY 5-Jun
Transformation Schedule
2:00 PM- Jacky E., Anita, Karuna, Grace, Marianna, Tony, Kavya, Oscar, Larab, Jason, Nioclet Oscar, Jason, Laraib =Make LB and LB plates
3:30 PM - Serena, Young, Vicky, Ashley, Renee, Brenden, Madalyn, Katherine V., Kevin, Melissa H.
4:00 PM - Alyssa, Priya, Ramiro, Jackie, Will I. (only doing check of plates and overnight culture)
THURSDAY 6-Jun
WELCH computer lab open - WELCH 2.144
10 am - 1 pm Protocol Target Discovery
Ashlee, Alyssa, Anita, Young, Katherine V., Manuel, Melissa, Nicolet, Priya, Ramiro , Vicky, William
2pm - 5 pm Protocol Target Discovery
Tony, Brendan, Jiaqi, Karuna, Renee, Kevin, Kavya, Madeline, Mariana, Serena, Oscar, Jason, Laraib, Serena, Grace, Jacky E,
- Check Transformation Efficiency
- Start overnight culture of your plasmid in DH5alpha in 80ml of LB (Maybe ???) - depends on timing
FRIDAY 7-JunThrow your overnight LB cultures into the fridge to 'stop' them so that you can spin them down later (a few hours)
10 am - 1 pm Protocol Target Discovery - WELCH 2.144
10 am - 'My First PCR'
Ashlee, Alyssa, Anita, Grace, Young, Katherine V., Manuel, Melissa, Priya, Ramiro , Vicky, William
Check Transformation Efficiency
Spin down LB flasks for doing MidiPrep later (store pellet in freezer)
2pm - 5 pm Protocol Target Discovery
2 pm - 'My First PCR'
Tony, Brendan, Jiaqi, Karuna, Renee, Kevin, Kavya, Madeline, Mariana, Serena, Oscar, Jason, Nicolet, Serena, Jacky E,
Input information about which culture of plasmid you have (or will) grow up.
http://vdsstream.wikispaces.com/Summer+Plasmids
Add content to YOUR research page
Research Pages 2013
Put a line and 'Week 1'
Below that add your 'result' image for the week with brief commentary
This page will run in reverse chronological order - i.e. Newest stuff on top
Order Overlap Primers for class
Journal article for next week is posted on Journal Club page.
SATURDAY 8-Jun
SUNDAY 9-Jun
Week 2
MONDAY 10-JunSafety training check.
Weekly Group Meeting @ 1:00 PM in GEA 100 (Gearing Hall)
Target Discovery selections
Protocol Primer Design OverlapAssembly
Order Overlap Primers for class
Transformations: Oscar, Jason, Laraib, Betty, Serena
Continue transformations
Do overnight cultures of plasmids in DH5alpha - check with Dr. B to see what/if you need to grow more
Input information about which culture of plasmid you have (or will) grow up.
http://vdsstream.wikispaces.com/Summer+Plasmids
TUESDAY 11-Jun
11:00 am Morning session of: Midi-prep plasmids -
- Tony, Ashlee, Alyssa, Anita, Grace, Young, Melissa, Priya, Vicky
1pm start agarose gels of last week's 1st PCR (for the people not doing MidiPrep - e.g. Katherine, Nicolet, Marianna, Kevin, Oscar, Jason, Serena, Kavya, Jacky)
PCR ? - Laraib, Betty
Submit to DNA sequencing - Laraib, Betty, Oscar, Jason
Nanodrop a plasmid - Laraib, Betty, Oscar, Jason
2:00 pm Afternoon session of: Midi-prep
-Brendan, Jiaqi, Karuna, Renee, Madeline, Manuel, Ramiro, Will
Do overnight cultures of plasmids - Oscar, Jason, Laraib, Betty
Make 500 ml of LB
New guys do Lab 1: Buffers & Solutions
On Your Targets Page:
Add Link to DNA Works output text fileAdd substrate cost, quantity, and catalog numbers (and supplier) for enzyme assay reagents
Add image of BRENDA reaction mechanism (screenshot it)
WEDNESDAY 12-Jun
11:30 am - Morning session of: Midi-prep plasmids - Katherine V, Nicolet, Marianna, Kevin, Oscar, Jason, Laraib, Betty, Serena, Kavya, Jacky
1pm start agarose gels of last week's 1st PCR
Fill out Travel Authorization Forms for field trips.
-- see GoogleDocs/Misc/TravelForms
-- print out and complete - put in Dr. B's box in lab (pigeon hole)
[ACTIONREQUIRED]: Will/Ramiro please put your DNAWorks output file into the "DNAWorks_Output" folder in vdsclass@gmail.com/Misc/Primers.
Grace/Young, Priya/Vicky, Jacky/Kavya, and Serena please upload your DNA oligonucleotide sequences onto the spreadsheet titled "PrimerPlateOrderTemplate_GDocs_Summer2013".
THURSDAY 13-Jun
Repeat 1st PCR if failed (why did it fail?)
Then:
ProtocolPCRforREcloningGREEN.xls OR ProtocolPCRforREcloningRED.xls
Analyze DNA Sequence Exercise as a group?
FRIDAY 14-Jun
11 am - 2pm Optional time to catch up on stuff.
- Daniel mentoring
Print Journal Club article for you and your friends
Make LB for next week (500ml per person)
Agarose gel checks
ProtocolPCRforREcloningGREEN.xls OR ProtocolPCRforREcloningRED.xls
Add content to YOUR research page on Wikispaces
Research Pages 2013
Lab Notebook check - should have everything up to date.
SATURDAY 15-Jun
-- sleep all day
SUNDAY 16-Jun
-- watch TV all day
WEEK 3
When not doing protein expression - can work on PCR's Red/Green, and then PCR pLIC sequencing,- ProtocolPCRforREcloningRED.xls OR ProtocolPCRforREcloningGREEN.xls (if fail, redo)
- ProtocolPCR_pLICsequencing.xls (if fail, redo)
Grow up your own batch of pNIC-Bsa4 (160ml) overnight and spin down to pellets the next day, save, and midi-prep later. (if fail, redo - because you need it for cloning)MONDAY 17-Jun
Those who need to update their Wikispaces page: Young, Karuna, Renee, Kevin, Madeline, Manuel, Nicolet, William
Verify your Safety training is all present.
1:00 - 4 people (Grace, Katherine, Young, Nicolet) start transformation of YopH and FtHAP into BL21(DE3) cells (instead of DH5alpha)
---- plasmids should be in 'Plasmids' box or the 'Final Plasmids' box
---- share your tube of competent bacterial cells BL21(DE3) with another groups - i.e. only use 1/2 of the tube for yours
Grace & Kavya - YopH in pNIC-Bsa4
Katherine & Jacky - FtHAP in pNIC-Bsa4
Young - YopH in pNIC-Bsa4
Nicolet & Kevin - FtHAP in pNIC-Bsa4
OTHERS:
Start one of the PCRs listed above
OR
Make LB for expression if you haven't already done so (500ml per person)
AND/OR
Prepare your flasks for overnight culture for protein expression (2x 50ml OR 2 x 100 ml of LB with Antibiotic) - Alyssa, Melissa, Vicky, Priya, Tony, Renee, Ramiro, Will
Label your flasks
AND/OR
Check DNA sequence of your MidiPrepped samples
-- once the plasmid is verified - move it to the regular 'plasmids' box (and but the other half in the top loading freezer in the 'plasmids' box)
-- Update plasmid info on the Google 'Plasmid Concentrations' spreadsheet:
---------https://docs.google.com/spreadsheet/ccc?key=0AoO2KqKh2q_-dENvNGJEdW9qdUxQZmFYeTVoeXA0SUE&usp=sharing
Weekly Group Meeting @ 1:00 PM CHANGED TO @ 3:00 PM in GEA 100 (Gearing Hall) - unless we can't get this room.
& Journal Club - see Journal Club page for paper
Daniel - create a 'Plasmids' box in the top loading freezer (already have one for the other freezer)
Analyze DNA sequence exercise (as a group) - in WEL computer lab
5: 00 - Overnight growth of pGBR22 2x 80 ml of LB, Midiprep tomorrow (Select 2 people) ? Betty & Laraib?
5:30 - start overnight cultures (only those people who are expressing tomorrow) (Alyssa, Melissa, Vicky, Priya, Tony, Renee, Ramiro, Will)
TUESDAY 18-Jun
10 am - Larger Scale Protein Expressions (work in pairs)
ProtocolProteinExpression_VDS_Summ13_v1.doc
USE THIS PROTOCOL TOO: ProtocolBacterialGrowthCurve_VDS_Summ13.xlsx
Alyssa (Not Mon or Wed) & Melissa - FabV in pNIC-Bsa4 PfDXR in pNic-BSSA4
Vicky & Priya - PfDXR in pNic-BSSA4
Tony & Renee (has class til 11:30) - FabI in pNIC-Bsa4PfDXR in pNic-BSSA4
Ramiro (Tues best, Monday ok, Not Wed, Thurs?) & Will - FrTUHP in pnic-BSA4 PfDXR in pNic-BSSA4
1:00 - everyone else come in to catch up on PCR's, agarose gel checks
Mentors (i.e. Michael) : During the break in expression
- make Resuspension buffer (~10ml per person) - see GDocs/Misc/Solutions&Dilutions or see protocol for protein expression
- make more 1 M IPTG
Transfer your transformation plate to 4degC fridge (label it correctly, and parafilm) - only these VDSers:
Grace & Kavya - YopH in pNIC-Bsa4
Katherine & Jacky - FtHAP in pNIC-Bsa4
Young - YopH in pNIC-Bsa4
Nicolet & Kevin - FtHAP in pNIC-Bsa4
5:30 - start overnight cultures (only those people who are expressing tomorrow) - Grace, Kavya, Katherine, Jacky, Madeline, Brendan, Kevin, Marianna, Serena, Jason, Oscar
WEDNESDAY 19-Jun
10 am - Larger Scale Protein Expressions (work in pairs)
ProtocolProteinExpression_VDS_Summ13_v1.doc
USE THIS PROTOCOL TOO: ProtocolBacterialGrowthCurve_VDS_Summ13.xlsx
Grace (Not Tuesday) & Kavya - (Kavya has every day til 11:30 am (could skip on any day but Monday) - prefers Wed) - YopH in pNIC-Bsa4
Katherine (but Tues afternoon class) & Jacky (not Mon - Tuesday is ok, Wednesday better) - FtHAP in pNIC-Bsa4
Madeline & Brendan - any day if advance notice..... - YopH in pNIC-Bsa4
Kevin (Kevin has class till 11:15) & Marianna - has class 8:30 - 11:30 (Tues is class all day) - FtHAP in pNIC-Bsa4
Serena FF @ 11 on Th - YopH in pNIC-Bsa4
Jason (10am free) & Oscar FtHap in pNIC-Bsa4
1:00 - people from yesterday - resuspend cell pellets in 10ml of buffer and store in conical tubes in -80degC
5:30 - start overnight cultures (only those people who are expressing tomorrow)
THURSDAY 20-Jun (Dr. B out of town) - Daniel and Co. are in charge
Firefighter Training ? - who is doing this....
Thurs @ 9:00 - 10: Karuna, Ashlee, Alyssa, Renee, Priya
Thurs @ 11- 12: Melissa, Tony, Oscar, Serena, Kavya, Anita, Marianna, Katherine
10 am - Larger Scale Protein Expressions (work in pairs)
ProtocolProteinExpression_VDS_Summ13_v1.doc
USE THIS PROTOCOL TOO: ProtocolBacterialGrowthCurve_VDS_Summ13.xlsx
Young - FtHAP in pNIC-Bsa4
Ashlee & Karuna (karuna has class til 11:30) - FabI in pNIC-Bsa4
Nicolet & Anita - YopH in pNIC-Bsa4
Betty & Laraib - (Laraib class 10-11:50 each day) - PfDXR in pNic-BSSA4
1:00 - people from yesterday - resuspend cell pellets in 10ml of buffer and store in conical tubes in -80degC
EVERYONE: complete and place your Release form and Medical Waiver in Dr. B's 'pigeon hole' (the one that says RESERVED over by the lab notebooks)
FRIDAY 21-Jun
(Dr. B out of town)
OFF
SATURDAY 22-Jun
-- engage in mental gymnastics
SUNDAY 23-Jun
-- invent something
PROTEIN GROUPS (from last week)
Group A
Alyssa & Melissa PfDXR in pNic-BSSA4
Vicky & Priya - PfDXR in pNic-BSSA4
Tony & Renee PfDXR in pNic-BSSA4
Ramiro & Will PfDXR in pNic-BSSA4
Group B
Grace & Kavya - YopH in pNIC-Bsa4
Madeline & Brendan YopH in pNIC-Bsa4
Kevin & Jacky- FtHAP in pNIC-Bsa4
Serena YopH in pNIC-Bsa4
Jason & Oscar FtHap in pNIC-Bsa4
Group C
Young - FtHAP in pNIC-Bsa4
Ashlee & Karuna FabI in pNIC-Bsa4
Nicolet & Anita YopH in pNIC-Bsa4
Betty & Laraib PfDXR in pNic-BSSA4
Katherine & Marianna FtHAP in pNIC-Bsa4
MONDAY 24-Jun
Weekly Group Meeting @ 1:00 PM in GEA 100 (Gearing Hall)
& Journal Club - see Journal Club page for paper
Analyze DNA sequence exercise (as a group) - in WEL computer lab
Check DNA sequence of your MidiPrepped samples
-- once the plasmid is verified - move it to the regular 'plasmids' box (and but the other half in the top loading freezer in the 'plasmids' box)
-- Update plasmid info on the Google 'Plasmid Concentrations' spreadsheet:
---------https://docs.google.com/spreadsheet/ccc?key=0AoO2KqKh2q_-dENvNGJEdW9qdUxQZmFYeTVoeXA0SUE&usp=sharing
EVERYONE: complete and place your Release form and Medical Waiver in Dr. B's 'pigeon hole' (the one that says RESERVED over by the lab notebooks)
TUESDAY 25-Jun
1:00Group A: Alyssa & Melissa PfDXR in pNic-BSSA4, Vicky & Priya - PfDXR in pNic-BSSA4, Tony & Renee PfDXR in pNic-BSSA4, Ramiro & Will PfDXR in pNic-BSSA4
Make Homemade Protein (SDS-PAGE) Gels
Sonication & spin down
Group B: Grace & Kavya - YopH in pNIC-Bsa4, , Madeline & Brendan YopH in pNIC-Bsa4, Kevin & Jacky- FtHAP in pNIC-Bsa4, Serena YopH in pNIC-Bsa4, Jason & Oscar FtHap in pNIC-Bsa4
Sonication & spin down (maybe on to purification?)
Group C: Young - FtHAP in pNIC-Bsa4, Ashlee & Karuna FabI in pNIC-Bsa4, Nicolet & Anita YopH in pNIC-Bsa4, Betty & Laraib PfDXR in pNic-BSSA4
Katherine & Marianna FtHAP in pNIC-Bsa4
RE digest (during incubation period - make gels)
5:00 - After RE digest, people from last Thursday's expression (Group C) - resuspend cell pellets in 10ml of buffer and store in conical tubes in -80degC (20-30 minutes)
WEDNESDAY 26-Jun
11:00 AM
Group A: Alyssa & Melissa PfDXR in pNic-BSSA4, Vicky & Priya - PfDXR in pNic-BSSA4, Tony & Renee PfDXR in pNic-BSSA4,
Sonication & Spin Down & Purification & run SDS-Page gels
1:00 PM
Group B: Grace & Kavya - YopH in pNIC-Bsa4, Madeline & Brendan YopH in pNIC-Bsa4, Kevin & Jacky- FtHAP in pNIC-Bsa4, Serena YopH in pNIC-Bsa4, Jason & Oscar FtHap in pNIC-Bsa4
RE digest (during incubation period - make gels) ------ Change to Purification
RE digest (during incubation period -
Make Homemade Protein (SDS-PAGE) Gels
Group C: Young - FtHAP in pNIC-Bsa4, Ashlee & Karuna FabI in pNIC-Bsa4, Nicolet & Anita YopH in pNIC-Bsa4, Betty & Laraib PfDXR in pNic-BSSA4, Katherine & Marianna FtHAP in pNIC-Bsa4,, and Ramiro & Will PfDXR in pNic-BSSA4
Sonication& Spin Down), purification
THURSDAY 27-Jun
10:00 AM
Group A: Alyssa & Melissa PfDXR in pNic-BSSA4, Vicky & Priya - PfDXR in pNic-BSSA4, Tony & Renee PfDXR in pNic-BSSA4, Ramiro & Will PfDXR in pNic-BSSA4
run gels, stain, destain
Group C: Young - FtHAP in pNIC-Bsa4, Ashlee & Karuna FabI in pNIC-Bsa4, Nicolet & Anita YopH in pNIC-Bsa4, Betty & Laraib PfDXR in pNic-BSSA4, Katherine & Marianna FtHAP in pNIC-Bsa4,
purification
For everyone else - Open time - come in to work on stuff (PCR's, gels, Wikispaces updating, Notebooks, etc.)
12:30 PM - NOTE TIME CHANGE - we need to be early enough to catch the bus.
EVERYONE: Gather together to get on bus to go to Pickle Research Campus for the TACC Tour (starts at 2pm there)
To get to PRC: take #3 Burnet/Manchaca bus.
4:00 - Dr. B meet with Vernier
Dr. B meet with Nicolet and Kevin about paper
Ramiro & Will PfDXR in pNic-BSSA4 - add Ni-NTA resing (only 0.5 ml) overnight
WEEK 4
FRIDAY 28-Jun -
11:00 AM
finish purifications, nanodrop, enter data to Google Spreadsheet (Protein Spectrophotometer Calcs)
Run gels (Protein gels), stain for 1 hour, destain overnight
Dry gels if already de-stained
Make protein gels (if we have enough combs)
RE digest if not doing protein stuff
EVERYONE:
SATURDAY 29-Jun
Play Golf
SUNDAY 30-Jun
Run a marathon
5th Week
MONDAY 1-Jul
Weekly Group Meeting @ 1:00 PM in GEA 100 (Gearing Hall)
& Journal Club - see Journal Club page for paper
Training is pretty much over
Target Assignments announced
see GDocs/Misc/ Targets/Target_Selections..
Design & Order 'Tail Primers': ProtocolPCR_PrimerDesign_for_pNIC-Bsa4CloningSumm13.doc
- the original, full gene sequences are in (be sure to use the Codon Optimized sequence!)
- also, don't use a file if it says 'BAD' on it.
GDOcs/Misc/Primers/DNAWorksOutputRe-run VS4 docking with Histidines protonated (Group 1)
TUESDAY 2-Jul
11:00 AM - mentors on duty
RE digest (if not done already)
Complete successful Practice PCRs (pGBR22 and then Red OR Green)
Finish up Tail Primer Design and Ordering: Nicolet, Tony, Renee, Kevin
Start Overlap PCRs
Re-run VS4 docking with Histidines protonated (Group 2)
Grow up your own pNIC-Bsa4, mdidiprep, sequence verify it, store in your own box
Cut your pNIC-Bsa4 to prepare the vector for cloning, save in -20degC in your box
WEDNESDAY 3-Jul
11:00 AM - mentors on duty
Dilute tail primers
submit pNIC-Bsa4 to sequencing, to verify plasmid & determine what gene is in the middle
THURSDAY 4-Jul
OFF - celebrate and shoot off some fireworks
FRIDAY 5-Jul
11:00 AM - mentors on duty
We'll have open lab time (with mentors)- for people to catch up on stuff but not an organized activity
??
1:00 Re-run VS4 docking with Histidines protonated (Group 2)
??
1:00 FPLC of PfDXR - for those that expressed it.
submit pNIC-Bsa4 to sequencing, to verify plasmid & determine what gene is in the middle
SATURDAY 6-Jul
Do people need more hours? - If so we can have lab times on Saturday. Let Dr. B know via email or in person (or carrier pigeon).
SUNDAY 7-Jul
Invent a new language
6th Week
PROTEN GROUPS FROM OTHER WEEK:
Group A: Alyssa & Melissa PfDXR in pNic-BSSA4, Vicky & Priya - PfDXR in pNic-BSSA4, Tony & Renee PfDXR in pNic-BSSA4, Ramiro & Will PfDXR in pNic-BSSA4
Group B: Grace & Kavya - YopH in pNIC-Bsa4, , Madeline & Brendan YopH in pNIC-Bsa4, Kevin & Jacky- FtHAP in pNIC-Bsa4, Serena YopH in pNIC-Bsa4, Jason & Oscar FtHap in pNIC-Bsa4
Group C: Young - FtHAP in pNIC-Bsa4, Ashlee FabI in pNIC-Bsa4, Nicolet & Anita YopH in pNIC-Bsa4, Betty & Laraib PfDXR in pNic-BSSA4
Katherine & Marianna FtHAP in pNIC-Bsa4
MONDAY 8-Jul
Weekly Group Meeting @ 1:00 PM in GEA 403 (Gearing Hall) -NOTE THE ROOM CHANGE!!!!!! Same building
& Journal Club - see Journal Club page for paperAfter class - PfDXR group spins down sample (combine together)
Alyssa & Melissa , Vicky & Priya, Tony & Renee PfDXR, Ramiro , Betty
4:00 - 5:15 - Naturally Obsessed documentary viewing in PAI 3.02 (with popcorn?)
TUESDAY 9-Jul
1:00 - FtHAP group spins down sample (combine together)
FtHAP group: Kevin & Jacky, Jason & Oscar, Katherine,
1 PM - FPLC for PfDXR group
Alyssa & Melissa , Vicky & Priya - PfDXR, Tony & Renee PfDXR, Ramiro PfDXR
2:00 Re-run VS4 docking with Histidines protonated (Group 2):
Melissa (?), Vicky, Priya, Tony, Renee , Ramiro,
Learn Virtual Screening:
Oscar, Jason, Stephanie, Brian, Gabe, Betty, Salil
4:00 - FPLC people collect fractions of protein and spin down again, Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
? Store in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC)
WEDNESDAY 10-Jul
1:00 - YopH group spins down sample (combine together)
YopH group: Grace & Kavya, Brendan, Serena, Nicolet, Young
1 PM - FPLC for FtHAP group
FtHAP group: Kevin & Jacky, Jason & Oscar, Katherine
2 - 4pm: Middle Schoolers (Shadow a Scientist) - Alyssa & Tony
4:00 - FPLC people collect fractions of protein and sping down again, Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
? Store in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC)
THURSDAY 11-Jul
1:30 or 2:00 PM - FPLC for YopH group
YopH group: Grace & Kavya, Serena, Young
1:00
PfDXR group - Alyssa & Melissa , Vicky & Priya, Tony & Renee, Ramiro - with ANDREW
YopH (i.e. PTP1b) group -Kevin, Jason & Oscar, Katherine - with DANIEL
Spin down FPLC'ed sample from the other day (to about 2 ml)
Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
Go get liquid nitrogen (with a mentor!)
Store in protein in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC)
1:00 - FtHAP group spins down sample (combine together)
3:00 Re-run VS4 docking with Histidines protonated (Group 3):
Grace, Young, Kavya, Ariel
Virtual Screening 1 (Part 2) - using GOLD: Oscar, Jason, Betty, Stephanie, Salil, Gabe, Brian
~ 4:00 - FPLC people collect fractions of protein and sping down again, Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
?
Wikispaces Research Pages that need to be updated: Brian C., Gabe H., Young, Jacky, Katherine, Kevin, Ramiro, - 070913
http://vdsstream.wikispaces.com/Research+Pages+2013
FRIDAY 12-Jul
Dr. B in a workshop all day
Daniel & Michael are in charge (scary!)
1:15 PM - TACC VisLab Tour (on campus in ACES building) - meet in our lab first and walk over
After TOUR:
YopH group: Grace & Kavya, Serena, Young
Spin down FPLC'ed sample from the other day (to about 2 ml)
Nanodrop it, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
Go get liquid nitrogen (with a mentor!)
Store in protein in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC)
1:00 - FtHAP group spins down sample (combine together)
EVERYONE:
SATURDAY 13-Jul
- bake a cake
SUNDAY 14-Jul
- cure cancer
ON YOUR OWN:
Move your Target page to top of Targets Page and add in missing information
- http://vdsstream.wikispaces.com/Targets
Update your Plasmid Concentrations to the Google Sheet if they have been verified by DNA sequencing:- https://docs.google.com/spreadsheet/ccc?key=0AoO2KqKh2q_-dENvNGJEdW9qdUxQZmFYeTVoeXA0SUE&usp=sharing
Grow up your own pNIC-Bsa4, midiprep, sequence verify it, store in your own boxafter submitting pNIC-Bsa4 to sequencing - determine what gene is in the middle
Cut your pNIC-Bsa4 to prepare the vector for cloning (one of the 1st steps in the cloning protocol), save in -20degC in your box
Continue Overlap PCR's
Begin cloning after successful PCR's
Target group meetings with Dr. B
Everyone make two (2) Kan+Sucrose LB Agar plates - we have lots (thank to Jacky, Oscar and Priya!) , but I want you to know how to make them.
-- make 20% (w/v) sucrose solution, sterile filter it ........ w/v = weight to volume
-- make LB agar plates as usual, autoclave
-- after cooled, add Kan and Sucrose (final conc of Sucrose should be 5% (w/v)) - NOTE: don't want to autoclave Kan or Sucrose
GOLD on your target - validation library
ICM run
VINA run, windows box only
MONDAY 15-Jul
Weekly Group Meeting @ 1:00 PM in GEA 403 (Gearing Hall) -NOTE THE ROOM CHANGE!!!!!! Same building
& Journal Club - see Journal Club page for paper2:30 - Session to update Wikispaces Target pages
Nanodrop protein, enter on Google spreadsheet (Protein Spectrophotometer Calcs)
Go get liquid nitrogen (with a mentor!)
Store in protein in Liquid Nitrogen (-80degC) and/or Glycerol (-20degC)
TUESDAY 16-Jul
WEDNESDAY 17-Jul
THURSDAY 18-Jul
People to update Target page:Nicolet, Grace & Young, Vicky & Kavya, Jacky & Kevin
People to update Wikispaces Research Pages: Grace, Jacky, Jason, Katherine, Nicolet, Ramiro,
Send your research presentation slides to Dr. B:Tony, Katherine V., Alyssa & Stephanie, Grace & Young
FRIDAY 19-Jul
Check your hours on the Hours Chart Be sure you have enough towards you 160.
EVERYONE:
SATURDAY 20-Jul
SUNDAY 21-Jul
MONDAY 22-Jul
Weekly Group Meeting @ 1:00 PM in GEA 403 (Gearing Hall)
& Journal Club - see Journal Club page for paperTUESDAY 23-Jul
Jacky, Kevin, Kavya, Vicky
grow up some pUC19
buy some Sma1 or HincII from ICMB Core Facility Store. Get NEB version.
Midi prep pUC19
make Xgal/IPTG plates with Ampicillin resistance
make some LB too (500ml)
Tues, July 23rd: Research Poster Design Workshop FAC 328 3-4 PM
- optional. Good preparation for the poster you will generate in the Spring for the Undergraduate Research Forum
WEDNESDAY 24-Jul
Send your research presentation slides to Dr. B: Nicolet, Ramiro, Jacky & Kevin, Melissa & Jason
THURSDAY 25-Jul
HSers (Brian, Gabe, Betty, Salil)
- work on Purchasing budget for reagents. In GoogleDrive/Misc/Purchasing
- examples of past orders are on second sheet in the file.
- copy and past a past order to the first sheet and then modify it to reflect what you used - Thx. - Dr. B
Then do VS3
Grace & Young - meet with Dr. B about journal paper
Send slides to Dr. B - Nicolet, Ramiro, Jacky & Kevin,
Journal Paper is uploaded:
http://vdsstream.wikispaces.com/Journal+Club+Summer+13
Social - 6 pm at Dr. B's
http://goo.gl/maps/M0KWQ
View Larger Map
FRIDAY 26-Jul
Check your hours on the Hours Chart Be sure you have enough towards you 160.
EVERYONE:
SATURDAY 27-Jul
SUNDAY 28-Jul